HMGB1 (high mobility group box 1) is a protein which has recently been rediscovered as an early or late inflammatory mediator distinct from the previously identified inflammatory cytokines (e.g., tumor necrosis factor and various interleukins) in diseases such as cerebral infarction, cerebral vasospasm, brain trauma, atherosclerosis, traumatic brain damage, sepsis, neuropathic pain and various types of arthritis, for which no therapeutic method has yet been sufficiently established; and HMGB1 is now receiving great attention because it can serve as a target of therapeutic and/or prophylactic methods for these diseases (Non-patent Document 1).
HMGB1 was discovered about 40 years ago as a protein showing high mobility during electrophoresis, which is among the non-histone proteins ubiquitously present in the cell nuclei of eukaryotic organisms and bound to chromatin. At first, this protein was referred to as HMG1 (high mobility group 1) as a member belonging to the high mobility group (HMG) protein family and was considered to play important roles in chromatin structure maintenance, transcriptional activity regulation and DNA repair, etc. Thereafter, it was rediscovered as a membrane binding protein (amphoterin) and further rediscovered again as an inflammatory mediator involved in various inflammatory diseases. In 2001, HMG1 was renamed HMGB1 as a result of reconsidering the nomenclature of the high mobility protein family.
HMGB1 protein is a 25 kDa protein composed of 215 amino acids rich in lysine residues and has an amino acid sequence which is very highly conserved among mammals. Its structure is composed of three domains, i.e., two DNA-binding domains called A-box (or box-A) and B-box (or box-B), and a carboxyl-terminal domain consisting only of aspartic acid and glutamic acid residues (also referred to as the C-terminal domain or acidic tail). The A-box and B-box are each composed of about 80 highly conserved amino acid residues, and are strongly positively charged. The B-box has a TLR4 (toll-like receptor 4)-binding domain and a RAGE (receptor for advanced glycation end products)-binding domain. Upon binding to TLR4, HMGB1 induces the secretion of inflammatory cytokines from macrophages/monocytes. Upon binding to RAGE, HMGB1 induces the growth, differentiation and migration of endothelial cells and other somatic cells (including tumor cells) and the expression of their cell surface proteins. The third domain, i.e., the carboxyl-terminal end has a structure consisting of a 30 amino acid sequence composed only of aspartic acid and glutamic acid residues and is excessively negatively charged. The amino acid sequence of this C-terminal segment is also known to be highly conserved among mammals, only with a few differences.
The HMGB1 protein was considered at first to have the functions of chromatin structure maintenance, transcriptional activity regulation, DNA repair and so on. However, particularly after 1999 when the HMGB1 protein was rediscovered as a late inflammatory mediator in sepsis by the research group of Tracey et al., discoveries have been made one after another, showing that the HMGB1 protein plays important roles in inflammatory cytokine cascades in various diseases. HMGB1 is not only localized in the nuclei of cells, but also migrates from the nuclei to the cytoplasm upon activation of macrophages and/or various cells of the immune system and is thereby secreted into the extracellular environment (active secretion). Alternatively, it has been elucidated that HMGB1 localized in the nuclei is rapidly released upon ischemia- or damage-induced cell necrosis or apoptosis (passive release). In recent years, HMGB1 or heat shock protein (HSP) or the like has been regarded as one of the endogenous damage-associated molecular patterns (DAMPs), which are released from damaged cells resulting from non-microbial causes (e.g., ischemia, trauma and the like). On the other hand, bacterial lipopolysaccharides (LPSs) and the like are referred to as pathogen-associated molecular patterns (PAMPs), which include various products of microbial origin. Receptors that recognize and respond to the latter patterns both on the cell surface and in the cytoplasm are referred to as pattern recognition receptors (PRRs), and their representative families include Toll-like receptors (TLRs). However, some members of the TLR family, particularly TLR2, TLR4 and TLR9 recognize and activate the above DAMPs. In particular, HMGB1 is known to activate TLR4 signaling and other events to induce inflammatory response, thereby resulting in enhanced TNFα secretion, etc. Moreover, as to RAGE, which is one of the HMGB1 receptors, it has been shown that RAGE-mediated transmission of inflammatory information plays an important role in amplification of this HMGB1-induced inflammatory response in diseases such as ischemia-induced brain disorder (Non-patent Document 2) and sepsis associated with bacterial infection (Non-patent Document 3), as a result of studies using RAGE knockout animals and/or studies using inhibitory peptides or specific antibodies against binding between RAGE and HMGB1. Namely, HMGB1 released into the extracellular environment acts as a strong inflammatory mediator via TLR4 or RAGE, etc., to further stimulate previously known inflammatory immune responses, so that HMGB1 may also be responsible for causing various serious diseases.
These diseases in which HMGB1 is involved (HMGB1-related diseases) are divided into two major groups, i.e., a group of diseases (e.g., septic shock) showing the extracellular secretion of HMGB1 resulting from microbial infection-induced immune responses, and a group of diseases (e.g., cerebral infarction) showing the extracellular release of HMGB1 caused by cell injury due to non-microbial causes. In the former group, TLR4 activation is induced, for example, by the action of bacterial components (e.g., bacterial lipopolysaccharides (LPSs)) produced upon infection. In response to this activation, monocytes, macrophages and other cells cause active secretion of HMGB1, which in turn acts as a late inflammatory mediator. HMGB1-related diseases in this context include sepsis, arthritis, atherosclerosis, various infections, and various immune diseases, etc. The latter group corresponds to cases where upon ischemia- or trauma-induced cell necrosis, HMGB1 having been localized in the nuclei is rapidly released into the extracellular environment within several hours (passive release) and thereby acts as an early inflammatory mediator to induce production of various inflammatory cytokines. Relevant diseases include cerebral infarction, traumatic brain injury, diseases due to ischemia during organ transplantation, myocardial infarction and so on.
In recent years, as to therapeutic or prophylactic methods for HMGB1-related diseases, reports have been made on studies searching for methods using an antibody against HMGB1 (Patent Documents 1, 2, 4 and 5), methods using a partial fragment of HMGB1 protein as an antagonist (Patent Documents 2 and 3), methods using an inhibitory compound against HMGB1 secretion (Non-patent Document 4) and so on. In particular, therapeutic methods using an antibody against HMGB1 in an animal model have been reported for the possibility of their application to sepsis (Patent Documents 1, 4 and 5), acute lung injury (Non-patent Document 5), connective tissue injury due to heat burn (Patent Document 2), arthritis (Patent Documents 4 and 5 and Non-patent Document 6), cerebral ischemia (Non-patent Document 7), amyloidosis (Patent Document 6), hepatopathy during intraportal islet transplantation (Non-patent Document 8) and neuropathic pain (Non-patent Document 9), etc. However, all of these studies have just been started as studies of therapeutic and prophylactic agents.
Under these circumstances, we have shown that rat-derived anti-HMGB1 monoclonal antibody is effective in animal models of cerebral infarction (Patent Document 8, Non-patent Documents 10 and 11), cerebral vasospasm (Patent Document 9), atherosclerosis (Patent Document 10 and Non-patent Document 12), traumatic brain damage (Patent Document 11 and Non-patent Document 13) and neuropathic pain (Non-patent Document 9). However, such a rat-derived antibody has a problem of immunogenicity and is difficult to use in humans.